[68Ga]Ga/[177Lu]Lu FAPI-04 from bench to bedside: first steps in CUDIM (Uruguay)

Introduction: Fibroblast activation protein (FAP) is a human enzyme that shows increased expression in the stroma of a variety of malignancies. It is also seen in more than 90% of activated stromal fibroblasts in all human carcinomas. Specific fibroblast activation protein (FAPI) inhibitors, labeled with different radionuclides, are being evaluated for the diagnosis of different tumors. Because FAP is highly expressed in cancer-associated epithelial fibroblasts, it can be used not only as a target for diagnosis, but it can also be used for therapeutic purposes.

Of all FAPI DOTA-FAPI-04 exhibited excellent properties, such as specific target, high-tumor uptake with low background, rapid clearance from blood, led the way for a clear diagnosis for the development of theragnostic studies based on FAPI derivatives. Our aim is to describe the labeling optimization of the derivative FAPI-04 with 68Ga and [¹⁷⁷Lu]Lu in order to have a theragnostic agent for its evaluation at the Uruguayan Center for Molecular Imaging (CUDIM).

Methods: FAPI-04 peptide was provided by MedChemExpress.

[⁶⁸Ga]68Ga was obtained from a generator (ITM). Activity levels ranged from 100 to 700 MBq. For labeling optimization, the following parameters were tested: i) FAPI-04 amount (10 to 75 mg); ii) pH (3.5 to 4.5); iii) incubation temperature (90 and 100 ºC); iv) incubation time (up to 20 min).

[¹⁷⁷Lu]LuCl₃ no-carried-added (130 to 830 MBq) was provided by ITM or ISOTOPIA. Precursor amount used was 1mg/37MBq. The following reaction medium were tested: i) 100 mL ascorbic acid 10% (w/v) and sodium acetate 0,32 M for pH adjustment (4,5 – 5,0); ii) 1mL fresh ascorbic acid buffer. The incubation temperature (95 and 100 ºC) and time (up to 20 min) were studied.

Radiochemical purity of both tracers was evaluated by ITLC and HPLC. In both cases, for the final purification, a C18 solid phase extraction cartridge (Sep-pak) was used. Labeling was performed in a GMP Radiopharmacy facility.

Results: [⁶⁸Ga]Ga-FAPI-04

Regarding to the precursor amount used, it was observed that to obtain a radiochemical yield greater than 85%, it is necessary to use at least 0.3 mg/MBq. The optimal pH was between 4.0 - 4.5 at 100 °C. The labelling yield performance did not increased after 10 min of heating.

Once the conditions were optimized manually, we proceeded to work on the semi-automatic synthesis platform (iQS® Ga-68 Fluidic Labeling Module). With this system, the complete eluate from the generator (4mL of 0.05M HCl) was used, so it was necessary to adjust the labeling conditions. The amount of precursor used was 25-75 mg with 890 mL of 0.32 M sodium acetate. The reaction was carried out for 10 min at 100 °C. The best conditions to obtain a labeling performance of 89% were with 25-30 mg of FAPI-04. The final product was obtained with radiochemical purity greater than 95%, which makes it optimal for clinical use as a PET/CT radiotracer.

[¹⁷⁷Lu]Lu-FAPI-04

For the labelling of FAPI-04 with [¹⁷⁷Lu]Lu, it was used in all cases a mass of 1 mg/37MBq and heating at 100 °C for 20 min. The best results were obtained with the ascorbic acid buffer adjusted to pH = 5.0 with NaOH (> 95%). The stability of the labelled peptide was verified up to 3 days. [¹⁷⁷Lu]Lu-colloid was not detected in any of the tests over time. The purification of the labelling was carried out under the same conditions described for [⁶⁸Ga]Ga-FAPI-04, obtaining a final radiochemical purity greater than 95%.

Conclusions: [¹⁷⁷Lu]Lu/[⁶⁸Ga]Ga-FAPI-04 were successfully synthesized in a GMP radiopharmacy facility. The development of novel theragnostic radiolabeled agents able to diagnose and treat cancer is being pursued with great interest. The identification of FAPI agents ([68Ga]Ga, [18F]F) opens a new way to diagnose and monitor cancer in a complementary way to [18F]FDG. At the same time giving patients the opportunity for therapy ([177Lu]Lu). It is our aim to look forward to translating it to the clinical setting and offer the benefit of these tracers.